The priority area of focus is "Preliminary, developmental, or pilot studies ancillary to ongoing research projects aimed at assessing the validity, utility, or added value of additional or novel methods to characterize the components, properties, or integrity of botanical/herbal/dietary supplement products." The parent grant is R01AT004314: Mechanisms of immunomodulatory and anti-tumor actions of Polysaccharide Krestin (PSK). In the parent grant, we plan to use the neu transgenic (tg) mouse, a model of ER-, HER2+ breast cancer, to study the immunomodulatory effects of protein-bound polysaccharide K (PSK), a commercially available mushroom extract. The specific aims of the parent grant are to: (1) determine whether PSK enhances activation and maturation of type I dendritic cells (DC1) in vivo and in vitro, (2) determine whether PSK can stimulate or augment an antigen specific immune response, (3) determine whether PSK can enhance tumor immunogenicity by up regulating immune and co-stimulatory molecules on the tumor as well as decreasing T regulatory cells in the tumor microenvironment. Results from the first year of the study have generated clear data demonstrating that PSK can enhance the function of DC and T cells. Furthermore, we found that the effect of PSK is mediated through specific activation of toll-like receptor 2 (TLR2). The study proposed in this revision will focus on the effect of PSK on gammadelta T cells, which was not included in the original application. Recent publications have shown that gammadelta T cells play an important role in antitumor immunity. It is reported that gammadelta T cells have high levels of TLR2 expression. Based on data generated from the current study, PSK activates TLR2, therefore PSK may affect the expansion and function of gammadelta T cells. PSK may also activate gammadelta T cells indirectly via activation of DC. Our preliminary experiment showed that including PSK in culture medium resulted in selective expansion of gammadelta T cells. Further investigation on this effect is very important for us to fully understand the anti-tumor mechanisms of PSK and to develop new PSK based immunotherapy. This is a novel direction and not included in the parent grant. The proposed study has three specific aims: (1) to determine the effect of PSK on gammadelta T cell expansion and lytic function in vitro;(2) to determine whether administration of PSK will expand gammadelta T cells in tumor-bearing neu-tg mice in vivo;and (3) to determine whether PSK stimulates gammadelta T cell proliferation via TLR2/DC interaction. Results from the proposed studies will expand the research scope of the parent grant which focuses on the effect of PSK adaptive immunity. PUBLIC HEALTH RELEVANCE: The proposed revision will expand the scope of the research aims of the parent grant by studying the effect of the mushroom extract polysaccharide krestin (PSK) on gammadelta T cells in a mouse model of breast cancer. The results generated from the revision will not only help us better understand the mechanisms of the antitumor effects of PSK, but also help us design novel PSK-based immunotherapy.